title: Correlative two-photon and light sheet microscopy.
abstract: Information processing inside the central nervous system takes place on multiple scales in both space and time. A single imaging technique can reveal only a small part of this complex machinery. To obtain a more comprehensive view of brain functionality, complementary approaches should be combined into a correlative framework. Here, we describe a method to integrate data from in vivo two-photon fluorescence imaging and ex vivo light sheet microscopy, taking advantage of blood vessels as reference chart. We show how the apical dendritic arbor of a single cortical pyramidal neuron imaged in living thy1-GFP-M mice can be found in the large-scale brain reconstruction obtained with light sheet microscopy. Starting from the apical portion, the whole pyramidal neuron can then be segmented. The correlative approach presented here allows contextualizing within a three-dimensional anatomic framework the neurons whose dynamics have been observed with high detail in vivo.
authors.affiliation: European Laboratory for Non-Linear Spectroscopy, University of Florence, Italy.
authors.forename: Ludovico
authors.initials: L
authors.lastname: Silvestri
authors.affiliation: European Laboratory for Non-Linear Spectroscopy, University of Florence, Italy.
authors.forename: Anna Letizia
authors.initials: AL
authors.lastname: Allegra Mascaro
authors.affiliation: European Laboratory for Non-Linear Spectroscopy, University of Florence, Italy.
authors.forename: Irene
authors.initials: I
authors.lastname: Costantini
authors.affiliation: National Institute of Optics, National Research Council, Sesto Fiorentino, Italy; European Laboratory for Non-Linear Spectroscopy, University of Florence, Italy.
authors.forename: Leonardo
authors.initials: L
authors.lastname: Sacconi
authors.affiliation: European Laboratory for Non-Linear Spectroscopy, University of Florence, Italy; National Institute of Optics, National Research Council, Sesto Fiorentino, Italy; Department of Physics and Astronomy, University of Florence, Italy; International Center for Computational Neurophotonics (ICON Foundation), Sesto Fiorentino, Italy. Electronic address: francesco.pavone@unifi.it.
authors.forename: Francesco Saverio
authors.initials: FS
authors.lastname: Pavone
mesh_terms: D000818:Animals; D003712:Dendrites / Q000648:ultrastructure*; D049452:Green Fluorescent Proteins / Q000096:biosynthesis; D051379:Mice; D008822:Mice, Transgenic; D018613:Microscopy, Confocal / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D013003:Somatosensory Cortex / Q000098:blood supply / Q000166:cytology*
chemical_list: D049452:Green Fluorescent Proteins
keywords: Brain imaging; Correlative microscopy; Light sheet microscopy; Two-photon fluorescence microscopy